WILD Helianthus annuus, A POTENTIAL SOURCE OF - REDUCED PALMITIC AND STEARIC FATTY ACIDS IN SUNFLOWER OIL

HELIA, 27, Nr. 40, p.p. 55-62, (2004)                                     UDC 633.854.78:665.12

  WILD Helianthus annuus, A POTENTIAL SOURCE OF
  REDUCED PALMITIC AND STEARIC FATTY ACIDS IN
                  SUNFLOWER OIL

          G.J. Seiler*

          U. S. Department of Agriculture, Agricultural Research Service,
          Northern Crop Science Laboratory, P. O. Box 5677, Fargo, ND 58105 USA

                                                                    Received: October 08, 2003
                                                                   Accepted: January 05, 2004

                                                 SUMMARY

                   The current trend in human diets is to decrease consumption of the satu-
             rated palmitic and stearic fatty acids. Healthy diets restricting not only total fat,
             but the saturated portion of that fat, would decrease blood serum cholesterol
             and the risk of coronary heart diseases. Edible vegetable oils are the principal
             source of fats in many diets. Sunflower oil, which is fifth in production among
             edible vegetable oils in the world, contains 65 g kg-1 saturated palmitic and 45
             g kg-1 saturated stearic acids. These concentrations are high compared to rape-
             seed oil with 40 g kg-1 palmitic and 20 g kg-1 stearic acids. A reduction of satu-
             rated fats in traditional sunflower oil would lead to a healthier edible oil. The
             objective of this preliminary study was to search the vast genetic diversity avail-
             able from wild Helianthus annuus, the closest relative of the cultivated sun-
             flower, for potential sources of reduced saturated fatty acids; less than 70 g kg-
             1
               combined palmitic and stearic fatty acids. Achenes of eighty-two populations
             of wild H. annuus were collected from the central Great Plains of the USA.
             Composited 20-achene samples from each population were analyzed for satu-
             rated fatty acids using organic base-catalyzed transesterification of fatty acid
             methyl esters and capillary gas chromatography. The average palmitic acid
             concentration ranged from 39 to 65 g kg-1 for the populations. Average stearic
             acid concentrations ranged from 19 to 37 g kg-1. Achene oil of one population
             of wild H. annuus from Holmquist, South Dakota, USA had a palmitic acid
             concentration averaging 39 g kg-1, while stearic acid averaged 19 g kg-1. The
             combined 58 g kg-1 palmitic and stearic acids is almost 50% lower than the
             present level of these fatty acids in sunflower oil. The level of saturated fatty
             acids observed in the population remained low when plants were grown in the
             greenhouse under uniform conditions. In the greenhouse, palmitic acid con-
             centration of this population averaged 40 g kg-1, while stearic acid averaged 19
             g kg-1. Crosses between this population and an inbred cultivated line produced
             F1 plants with an achene oil that averaged 39 g kg-1 palmitic and 21 g kg-1
             stearic acid. In comparison, the inbred cultivated parent averaged 61 g kg-1
             palmitic and 51 g kg-1 stearic acid. F2 plants produced achene oil that aver-
             aged 45 g kg-1 palmitic and 23 g kg-1 stearic acid, for a total of 68 g kg-1. When

  * Corresponding author, Phone: (1) 701-239-1380, Fax: (1) 701-239-1346,
    e-mail: seilerg@fargo.ars.usda.gov

56                                                HELIA, 27, Nr. 40, p.p. 55-62, (2004)

             F1 plants were backcrossed to the cultivated inbred, BC1F1 plants produced an
             achene oil that averaged 45 g kg-1 palmitic and 26 g kg-1 stearic acid for a total
             of 71 g kg-1. In comparison, the inbred cultivated parent averaged 65 g kg-1
             palmitic and 42 g kg-1 stearic acid, for a total of 107 g kg-1. Preliminary infor-
             mation indicates that palmitic and stearic fatty acids in sunflower oil can be
             reduced by introducing genes from a wild annual population into cultivated
             sunflower. Further research will be needed to determine the inheritance of
             these fatty acids. Other agronomic traits will also have to be monitored during
             introgression of the fatty acids genes.

             Key words:    palmitic acid, stearic acid, saturated fatty acids, H. annuus, wild
                           species

                                  INTRODUCTION
     In recent years consumers have become concerned about the consumption of
saturated fats in their diets. High levels of saturated fat consumption may contrib-
ute to increased blood serum cholesterol which in turn increases the risk of coro-
nary heart disease (Mensink et al., 1994; Willett, 1994). Prompted by nutritional
recommendations to consume fats lower in saturates and food manufacturers'
interest in reducing the use of hydrogenated oil, food processors are interested in
sunflower oil with specific fatty acid profiles (Fitch-Haumann, 1994). Vegetable oils
are the principle source of fats in many diets. Compared with many edible vegetable
oils, the saturated fatty acid concentration of 120 g kg-1 in sunflower (Helianthus
annuus L.) oil is considered moderate, with the principal saturated fatty acids
being palmitic (65 g kg-1) and stearic (45 g kg-1) acids. Canola (Brassica napus L.)
oil with only 60 g kg-1 is considered low in saturated fats. A reduction of saturated
fatty acids in sunflower oil to the 60 to 80 g kg-1 level would enhance the acceptabil-
ity of sunflower oil.
     The genus Helianthus contains 51 species, 37 perennial and 14 annual (Schill-
ing and Heiser, 1981). Wild Helianthus species have been used to improve the eco-
nomic and agronomic characteristics of cultivated sunflower (Seiler, 1992; Seiler
and Rieseberg, 1997). Considerable emphasis has been placed on oil concentration
and fatty acid composition of the oil. Interest has centered on the enhancement of
the linoleic or oleic fatty acids, and the reduction of saturated palmitic and stearic
fatty acids. Wild sunflower species provide a resource for improving fatty acid com-
position in cultivated sunflower (Dorrell and Whelan, 1978; Thompson et al., 1981;
Seiler, 1985, 1994). Potential sources with lower saturated palmitic and stearic
acids from the wild species have been identified (Seiler, 1994, 1998), but their sta-
bility and transfer into cultivated sunflower have not been documented.
     This study evaluated populations of wild annual H. annuus, the closest relative
of cultivated sunflower, for lower palmitic and stearic acids, to determine their sta-
bility, and explored the possibility of introgressing the lower saturated fatty acid
genes into cultivated sunflower.

HELIA, 27, Nr. 40, p.p. 55-62, (2004)                                                   57

                         MATERIALS AND METHODS
     Achenes of eighty-two populations of H. annuus were collected from the central
Great Plains of the USA (Seiler, 1994). Achenes were stored at 5°C and low humid-
ity (

58                                                  HELIA, 27, Nr. 40, p.p. 55-62, (2004)

average palmitic acid concentration with 41 g kg-1, while the highest observed was
from Kansas with 56 g kg-1.
    A single population of wild H. annuus (ANN-2229, PI 586886) from Holmquist,
South Dakota, USA, had the lowest total saturated fatty acid concentration of 58 g
kg-1 with a palmitic acid concentration of 39 g kg-1, and a stearic acid concentration
with 19 g kg-1. This concentration is 50% lower than that generally observed in oil
of cultivated sunflower. This population was chosen to determine the stability of the
reduced saturated fatty acid trait to see if the genes controlling these acids were
dominant, and if they could be transferred into cultivated sunflower.
    Progeny of the ANN-2229 population were grown in the greenhouse at 22-25°C
and 16 hours of daylight. The plants were sib-pollinated. The saturated fatty acids
in achene oil of the population were very similar to the levels observed in the origi-
nal population (Table 1).
Table 1: Comparison of palmitic and stearic fatty acids from plants of wild H. annuus (ANN-
         2229) from the original population and plants grown in a common environment in
         the greenhouse
Environment                         Palmitic acid         Stearic acid         Total
                                        g kg-1              g kg-1            g kg-1
ANN-2229 (Original)                      47                   18                65
ANN-2229 (Greenhouse)                    48                   16                64
Cultivated NMS HA89 (GH)                 65                   44               109

     A standard cultivated line, fertile NMS HA89, was also grown as a control in the
greenhouse and had a palmitic acid concentration of 65 g kg-1 and a stearic acid
concentration of 44 g kg-1 for a total of 109 g kg-1. The low levels of saturated fatty
acids observed in the original population appear to be stable, indicating that the
reduced levels of palmitic and stearic acids have a genetic base and the potential to
be introgressed into cultivated sunflower.
     The F1 achenes produced in the field had an average palmitic acid concentra-
tion of 39 g kg-1 and a stearic acid concentration of 21 g kg-1 in the oil. These values
were the average of 10 F1 plants.
     The cultivated inbred line NMS HA 89 used to produce F1 hybrids averaged 61
g kg-1 palmitic and 51 g kg-1 stearic fatty acids. F1 plants were self-pollinated in the
field to produce F2 achenes. Achene oil of field grown F2 plants averaged 41 g kg-1
palmitic acid and 18 g kg-1 stearic acid totaling 59 g kg-1 of saturated fatty acids.
The averages were based on 10 individual plants.
     F1 plants were backcrossed in the field with cultivated NMS HA89 as the female
to produce BC1F1 achenes. Achene oil of the BC1F1 plants averaged 38 g kg-1 pal-
mitic acid and 19 g kg-1 stearic acid. These values were based on 10 plants from
one backcross family. In comparison, the cultivated NMS HA 89 line averaged 65 g
kg-1 of palmitic acid and 42 g kg-1 of stearic acid.

HELIA, 27, Nr. 40, p.p. 55-62, (2004)                                                        59

                                     CONCLUSIONS
     Preliminary results indicate that palmitic and stearic acid levels in sunflower oil
can be lowered by the introgression of genes from a population of the closest wild
relative of the cultivated crop. The genes appear to be relatively stable after transfer.
Further research will be needed to determine the inheritance of the genes control-
ling palmitic and stearic fatty acids. Acceptable agronomic traits will also have to be
bred into the lines during the introduction of the genes into cultivated sunflower.

                                      REFERENCES

Dorrell, D.G., Whelan, E.D.P., 1978. Chemical and morphological characterization of seeds of
       some sunflower species. Crop Sci. 18: 969-971.
Fitch-Haumann, B., 1994. Modification of oil may be the key to sunflower's future. INFORM-
       International News on Fats, Oil, and Related Materials, 5(1): 1198-1210.
Mensink, R.P., Temple, E.H.M., Hornstra, G., 1994. Dietary saturated and trans fatty acids and
       lipoprotein metabolism. Ann. Med. 26: 461-464.
Metcalfe, L.D., Wang, C.N., 1981. Rapid preparation of fatty acid methyl esters using organic
       base catalyzed transesterification. J. Chrom. Sci. 19: 530-535.
Schilling, E.E., Heiser, C.B., Jr., 1981. Infrageneric classification of Helianthus (Compositae).
       Taxon 30: 393-403.
Seiler, G. J., 1985. Evaluation of seeds of sunflower species for several chemical and morpho-
       logical characteristics. Crop Sci. 25: 183-187.
Seiler, G.J., 1992. Utilization of wild sunflower species for the improvement of cultivated
       sunflower. Field Crops. Res. 30: 195-230.
Seiler, G.J., 1994. Oil concentration and fatty acid composition of achenes of North American
       Helianthus species. Econ. Bot. 48: 272-279.
Seiler, G.J., 1998. The potential use of wild Helianthus species for selection of low saturated
       fatty acids in sunflower oil. In: A.M de Ron (ed.), International Symposium on Breeding
       of Protein and Oil Crops, EUCARPIA, Pontevedra, Spain, pp. 109-110.
Seiler, G. J., Rieseberg, L.R., 1997. Systematics, origin, and germplasm resources of the wild
       and domesticated sunflower. In: A. Schneiter (ed.) Sunflower Technology and Production.
       Crop Science Society of America, Madison, WI, pp. 21-65.
Thompson, T.E., Zimmerman, D.C., Rogers, C.E., 1981. Wild Helianthus as a genetic resource.
       Field Crops Res. 4: 333-343.
Willett, W.C., 1994. Diet and health: What should we eat? Science 264: 532-537.

               Helianthus annuus SILVESTRE COMO UNA FUENTE
               POTENCIAL DE CONTENIDO DISMINUIDO DEL ÁCIDO
               PALMÍTICO Y ESTEÁRICO EN EL ACEITE DE GIRASOL
                                                  RESUMEN

                     Actualmente en la alimentación humana está presente la tendencia hacia
               un reducido consumo del ácido palmítico y esteárico saturados. Con una ali-
               mentación sana, en la cual está limitado no sólo el contenido total de grasas,
               sino también el contenido de la parte saturada de dicha grasa, se disminuiría
               el contenido de colesterol en el suero sanguíneo y el riesgo de las enferme-
               dades cardíacas coronarias. Los aceites comestibles de origen vegetal, son la
               principal fuente de grasas en muchas dietas. El aceite de girasol, que entre los
               aceites vegetales comestibles, ocupa el quinto lugar según las cantidades pro-
               ducidas en el mundo, contiene 65 g kg-1 del ácido palmítico saturado y 45 g kg-
               1
                  del ácido esteárico saturado. Estas concentraciones son altas en com-
               paración con el aceite de colza, que contiene 40 g kg-1 de ácido palmítico, y 20

60                                        HELIA, 27, Nr. 40, p.p. 55-62, (2004)

     g kg-1 del ácido esteárico. Reduciendo el contenido de grasas saturadas en el
     aceite de girasol estándar, se obtendría aceite comestible más sano. El objetivo
     de esta investigación preliminar era, en la enorme divergencia genética que
     existe en el girasol silvestre (Helianthus annuus), el más cercano familiar del
     girasol cultivado, de buscar fuentes potenciales del reducido contenido de áci-
     dos grasos, es decir, las fuentes con menos de 70 g kg-1 del ácido palmítico y
     esteárico juntas. Los aquenios de 82 poblaciones de girasol silvestre, se rec-
     olectaron en la región de Great Plains en lso E.E.U.U. Las muestras compues-
     tas de 20 aquenios de cada población, fueron analizados para el contenido de
     los ácidos grasos saturados, mediante transesterificación de metilo ester de
     ácidos grasos catalizados con las bases orgánicas y cromatografía capilar de
     gases. La concentración promedia del ácido palmítico en las poblaciones inves-
     tigadas varía entre 39 y 65 g kg-1. La concentración promedia del ácido
     esteárico, era entre 19 y 37 g kg-1. En una población de girasol silvestre, del
     lugar llamado Holmquist en Dakota del Sur (E.E.U.U.), la concentración pro-
     media del ácido palmítico en el aceite de aquenios, era 39 g kg-1, y el contenido
     promedio del ácido esteárico, era 19 g kg-1. La suma concentración del ácido
     palmítico y del esteárico de 58 g kg-1 es más baja por 50% del contenido actual
     de esos ácidos grasos en el aceite de girasol. El contenido de los ácidos grasos
     saturados en esta población se ha quedado bajo, también cuando se cultivaban
     las plantas en el invernadero, en las condiciones uniformes. En el invernadero,
     la concentración promedia del ácido palmítico en esa población, era 40 g kg-1 y
     del esteárico, 19 g kg-1. Con cruzamientos entre esta población y una línea con-
     sanguínea del girasol cultivado, se obtuvieron las plantas F1 con el aceite de
     aquenios que contiene, en promedio, 39 g kg-1 de ácido palmítico, y 21 g kg-1
     del ácido esteárico. Por comparación, la línea consanguínea utilizada como
     padre, en un promedio tiene 61 g kg-1 de ácido palmítico y 51 g kg-1 del ácido
     esteárico. Las plantas F2 tenían el aceite de aquenios, con el promedio de 45 g
     kg-1 de ácido palmítico y 23 g kg-1 del ácido esteárico, lo que en suma,
     asciende a 68 g kg-1. Una vez efectuados los cruzamientos reversibles entre las
     plantas de F1 y dicha línea consanguínea, las plantas BC1F1 dieron aceite de
     aquenio, que en un promedio tiene 45 g kg-1 de ácido palmítico y 26 g kg-1 de
     ácido esteárico, o sea, en total, 71 g kg-1, mientras que la línea consanguínea
     utilizada como padre, tiene en un promedio, 65 g kg-1 de ácido palmítico y 42
     g kg-1 de ácido esteárico, o sea, en total, 107 g kg-1. Las informaciones prelim-
     inares sugieren que el contenido del ácido palmítico y el esteárico en el aceite
     de girasol, puede reducirse, introduciendo los genes de las poblaciones silves-
     tres anuales en el girasol cultivado. Se necesitan investigaciones adicionales,
     para determinar la forma de herencia de esos ácidos grasos. También tendrán
     que seguirse las demás propiedades agronómicas, durante la introgresión del
     gen para estos ácidos grasos.

     ESPECE SAUVAGE Helianthus annuus, COMME SOURCE
     POTENTIELLE DE RÉDUCTION DE GRAS ACIDES
     PALMITIQUES AND STÉARIQUES DANS L’HUILE DE
     TOURNESOL
                                        RESUME

          Actuellement dans le régime de nourriture humaine, une tendance est
     présente vers la réduction de consomation d’acides saturés, palmitiques et
     stéariques. La nourriture saine limite, non seulement le total de matière
     grasse, mais le contenu saturé de matière grasse afin de diminuer le contenu
     de cholestérol dans le sérum sanguin et le danger de maladies coronaires, car-
     diaques. Les huiles comestibles d’origine végétale sont la source principale de
     régimes diététiques. L’huile de tournesol, qui est au cinquième rang dans la
     production d’huiles comestible, contient 65g kg-1 d’acide saturé palmitique et

HELIA, 27, Nr. 40, p.p. 55-62, (2004)                                                       61

             45g kg-1 d’acide saturé stéarique. Ces concentrations sont assez élevées en
             comparaison de l’huile de colza qui contient 40g kg-1d’acide palmitique et 20g
             kg-1 d’acide stéarique. Par la réduction de contenu d’acide gras saturé dans
             l’huile standard de tournesol, une huile comestible saine pourrait être obtenue.
             Le but de cette étude préliminaire pourrait servir de trouver les sources poten-
             tielles de contenu faible d’acides gras, c’est-à-dire les sources d’acides palmi-
             tiques et stéariques moins de 70g kg-1au total dans l’énorme divergence
             génétique d’espèces sauvages de Helianthus annuus, le parent le plus proche
             du tournesol cultivé. Les achènes de 82 populations de tournesol sauvage sont
             recueillis dans la région de Great Plains aux Etats Unis. Les échantillons com-
             posés de 20 achènes de chaque population sont analysés pour déterminer le
             contenu d’acides gras saturés au moyen de trans-estérification d’acides gras du
             méthyle-ester, catalysés par les bases organiques et par la chromatographie
             capillaire gazeuse. La concentration moyenne d’acide palmitique de la popula-
             tion examinée était entre 39 et 65 g kg-1. La concentration moyenne d’acide
             stéarique était entre 19 et 37 g kg-1. Chez une population du tournesol sauvage
             de Holmquist en Dakota de Sud aux Etats Unis, une concentration moyenne
             d’acide palmitique dans l’huile d’achène, était enregistrée de 39 g kg-1, tandis
             que le contenu d’acide stéarique était de 19 g kg-1. La somme totale de concen-
             trations d’acides palmitiques et stéariques de 58 g kg-1, est inférieure de 50%
             du contenu actuel de tous les acides gras dans l’huile de tournesol. Le contenu
             d’acides gras saturés de cette population est resté faible même si les plantes
             sont cultivées dans les conditions uniformes d’orangerie. Dans les conditions
             d’orangerie, la concentration moyenne d’acide palmitique de cette population
             était de 40 g kg-1, tandis que la valeur d’acides stéarique était de 19 g kg-1. Par
             le croisement entre cette population et une ligne de tournesol cultivé, les plan-
             tes de F1 sont obtenues d’une huile d’achène ayant, en moyenne, les acides
             palmitiques de 39 g kg-1 et acides stéariques de 21 g kg-1. Comparant la ligne
             cultivée, utilisée comme parent qui contient, en moyenne, d’acides palmitiques
             de 61 g kg-1 et acides stéariques de 51 g kg-1avec les plantes de F1qui pos-
             sèdent l’huile d’achène, en moyenne, d’un contenu d’acides palmitiques de 45 g
             kg-1 et acides stéariques de 23 g kg-1, dont la somme totale est de 68 g kg-1.
             Quand les croisements réciproques sont réalisés entre les plantes de F1 et les
             lignes mentionnées, les plantes BC1F1 ont donné l’huile d’achène, en moyenne,
             d’acides palmitiques de 45 g kg-1 et acides stéariques de 26 g kg-1, c’est-à-dire
             la somme totale est de 71 g kg-1, tandis que la ligne cultivée, utilisée comme
             parent contient, en moyenne, d’acides palmitiques de 65 g kg-1 et acides stéar-
             iques de 42 g kg-1, c’est-à-dire la somme totale est de 107 g kg-1. Les informa-
             tions préliminaires indiquent que le contenu d’acides palmitiques et stéariques
             dans l’huile de tournesol pourrait être diminué par introduction du gène de
             sauvages populations annuelles aux lignes cultivées. Les recherches
             postérieures sont nécessaires afin de déterminer le procédé d’hérédité d’acides
             gras. Les autres traits agronomiques devraient être contrôlés pendant l’intro-
             gression de gènes d’acides gras

62   HELIA, 27, Nr. 40, p.p. 55-62, (2004)